The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation
Summary: ATP-powered unfoldases containing D1 and D2 AAA+ rings play important roles in protein homeostasis, but uncertainty about the function of each ring remains. Here we use single-molecule optical tweezers to assay mechanical unfolding and translocation by a variant of the ClpAP protease contai...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2020-02-01
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| Series: | Cell Reports |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2211124720301546 |
| _version_ | 1828169825860452352 |
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| author | Hema Chandra Kotamarthi Robert T. Sauer Tania A. Baker |
| author_facet | Hema Chandra Kotamarthi Robert T. Sauer Tania A. Baker |
| author_sort | Hema Chandra Kotamarthi |
| collection | DOAJ |
| description | Summary: ATP-powered unfoldases containing D1 and D2 AAA+ rings play important roles in protein homeostasis, but uncertainty about the function of each ring remains. Here we use single-molecule optical tweezers to assay mechanical unfolding and translocation by a variant of the ClpAP protease containing an ATPase-inactive D1 ring. This variant displays substantial mechanical defects in both unfolding and translocation of protein substrates. Notably, when D1 is hydrolytically inactive, ClpAP often stalls for times as long as minutes, and the substrate can back-slip through the enzyme when ATP concentrations are low. The inactive D1 variant also has more difficulty traveling in the N-to-C direction on a polypeptide track than it does moving in a C-to-N direction. These results indicate that D1 normally functions as an auxiliary/regulatory motor to promote uninterrupted enzyme advancement that is fueled largely by the D2 ring. : Kotamarthi et al. employ optical tweezers to examine the individual rings in a double-ring AAA+ unfoldase. ClpAP inactivated in the D1 ring is partially defective in unfolding and pauses frequently during translocation. Thus, D1 functions as a booster motor, thereby assisting the more powerful unfoldase motor of the D2 ring. Keywords: AAA+ proteases, ATP motors, single-molecule force spectroscopy, protein unfolding, protein translocation, translocation pauses, Hsp104/ClpB, ClpAP, optical tweezers, protein degradation |
| first_indexed | 2024-04-12T02:57:20Z |
| format | Article |
| id | doaj.art-8b5c85d192f14b438dfb03c16046b56e |
| institution | Directory Open Access Journal |
| issn | 2211-1247 |
| language | English |
| last_indexed | 2024-04-12T02:57:20Z |
| publishDate | 2020-02-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Cell Reports |
| spelling | doaj.art-8b5c85d192f14b438dfb03c16046b56e2022-12-22T03:50:45ZengElsevierCell Reports2211-12472020-02-0130826442654.e3The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and TranslocationHema Chandra Kotamarthi0Robert T. Sauer1Tania A. Baker2Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA; Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA, USADepartment of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA; Corresponding authorDepartment of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA; Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA, USA; Corresponding authorSummary: ATP-powered unfoldases containing D1 and D2 AAA+ rings play important roles in protein homeostasis, but uncertainty about the function of each ring remains. Here we use single-molecule optical tweezers to assay mechanical unfolding and translocation by a variant of the ClpAP protease containing an ATPase-inactive D1 ring. This variant displays substantial mechanical defects in both unfolding and translocation of protein substrates. Notably, when D1 is hydrolytically inactive, ClpAP often stalls for times as long as minutes, and the substrate can back-slip through the enzyme when ATP concentrations are low. The inactive D1 variant also has more difficulty traveling in the N-to-C direction on a polypeptide track than it does moving in a C-to-N direction. These results indicate that D1 normally functions as an auxiliary/regulatory motor to promote uninterrupted enzyme advancement that is fueled largely by the D2 ring. : Kotamarthi et al. employ optical tweezers to examine the individual rings in a double-ring AAA+ unfoldase. ClpAP inactivated in the D1 ring is partially defective in unfolding and pauses frequently during translocation. Thus, D1 functions as a booster motor, thereby assisting the more powerful unfoldase motor of the D2 ring. Keywords: AAA+ proteases, ATP motors, single-molecule force spectroscopy, protein unfolding, protein translocation, translocation pauses, Hsp104/ClpB, ClpAP, optical tweezers, protein degradationhttp://www.sciencedirect.com/science/article/pii/S2211124720301546 |
| spellingShingle | Hema Chandra Kotamarthi Robert T. Sauer Tania A. Baker The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation Cell Reports |
| title | The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation |
| title_full | The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation |
| title_fullStr | The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation |
| title_full_unstemmed | The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation |
| title_short | The Non-dominant AAA+ Ring in the ClpAP Protease Functions as an Anti-stalling Motor to Accelerate Protein Unfolding and Translocation |
| title_sort | non dominant aaa ring in the clpap protease functions as an anti stalling motor to accelerate protein unfolding and translocation |
| url | http://www.sciencedirect.com/science/article/pii/S2211124720301546 |
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